HPRT YEAST ARTIFICIAL CHROMOSOME TRANSFER INTO HUMAN-CELLS BY 4 METHODS AND AN INVOLVEMENT OF HOMOLOGOUS RECOMBINATION

The transfer of a yeast artificial chromosome (YAC) into mouse and hum an cell lines was effected by four methods, and the efficiency and int egrity of the incorporated YAC DNA were compared. A 500 kb YAC contain ing the human hypoxanthine-guanine phosphoribosyl transferase (HPRT) g ene was transferred more efficiently by polyethylene glycol-mediated f usion than by lipofection, electrofusion, or electroporation. Southern blot analysis demonstrated that PEG fusion lines yielded fragments of the size of the original YAC clone, whereas lipofection and electropo ration did not. Two of 53 fusion lines showed 6-thioguanine resistance and confirmatory disruption of the HPRT gene in the YAC DNA, that the YAC DNA was integrated by homologous recombination with the HPRT gene region. (C) 1994 Academic Press, Inc.